Haemophilus influenzae (NTHi) is a major pathogen which causes otitis media in children. Lipooligosaccharide (LOS), one of the most important surface antigens in NTHi, is immunogenic and a potential target of bactericidal antibodies during Haemophilus otitis media. We are producing a LOS-derived OS-protein conjugate as a possible vaccine candidate to induce a long term immune response for the prevention of NTHi otitis media in young children. Three NTHi strains were selected as vaccine candidates based on their different LOS molecular sizes and their immunogenicity in mice. The mouse antisera showed bactericidal activities on both homologous and heterologous strains. To prepare high yields of LOSs from NTHi, several growth conditions and isolation procedures were examined. The best procedure for the purification of the LOS appears to be the combined methods of Westphel et al. (1965) and Gu & Tsai (1991). The resulting yield of this procedure is about a two-fold increase in the LOS. We are currently detoxifying the LOSs and producing oligosaccharide-protein conjugates from 3 LOSs by using different chemical methods to preserve the antigenicity of the native LOSs. In order to characterize the specific epitomes on NTHi for bactericidal antibody response, twenty-one mouse monoclonal antibodies (MAbs) against strain 9274 were prepared. The isotypes, specificity and functions of these MAbs are being characterized. The preliminary results indicate that two of these MAbs are IgG3 and others are IgM. Two of these MAbs are against outer membrane proteins and others are against LOS. Interestingly, five of the MAbs which are against the LOS showed bactericidal activities.